【摘要】： 目的: 采用單細胞染色體制備技術(shù),分析人類常規(guī)體外受精與胚胎移植中(in-vitro fertilization and embryo transfer, IVF-ET )異常受精卵及異常受精卵來源胚胎的染色體組成。 方法: 加精后24～72小時收集異常受精卵75枚、異常受精卵來源的胚胎277枚,其中IVF來源的三原核(tripronuclear, 3PN)受精卵71枚、2細胞胚胎38枚、2細胞胚胎239枚,卵胞漿內(nèi)單精子顯微注射(intracytoplasmic sperm injection,ICSI)來源的單原核(monopronucleus, 1PN)胚胎4枚。分別與秋水仙素(終濃度0.10 ug/ml)共培養(yǎng)14～20小時,Tyrode’s液去除透明帶,胚胎需分散成卵裂球,0.9%枸櫞酸鈉液低滲20～30分鐘后梯度固定,胰酶顯帶、Giemsa染色,美國維士染色體核型自動分析系統(tǒng)(Auto-System K)分析核型并照相。 結(jié)果: (1) IVF-3PN受精卵及胚胎的染色體制備:3PN受精卵的有絲分裂指數(shù)及染色體制備成功率最高,分別為67.6% (48/71)、52.1% (37/71)。2細胞胚胎的有絲分裂指數(shù)及染色體制備成功率分別為38.2%(29/76)、31.2%(24/76)。2細胞胚胎的有絲分裂指數(shù)及染色體制備成功率最低,分別為16.2%(149/920)、8.2%(75/920)。 (2) 3PN受精卵及胚胎的染色體組成:3PN受精卵的三倍體率最高,為75.7%(28/37),2細胞胚胎為70.8%(17/24),2細胞胚胎染色體核型紊亂,多為近二倍體(56%)、近單倍體(32%),三倍體率僅為9.3%(7/75)。 (3) ICSI-1PN胚胎的染色體組成:4枚ICSI-1PN胚胎分散后共獲29枚細胞,其中6細胞(來自于3枚胚胎)可染色體計數(shù),顯示均為單倍體(5細胞為23條染色體,1細胞為25條染色體)。 結(jié)論: (1)傳統(tǒng)染色體制備方法可對受精卵及胚胎行全套染色體檢測,但技術(shù)操作難度大、染色體制備成功率低,該方法尚需進一步改進。 (2) IVF-3PN受精卵及-2細胞胚胎多為三倍體,2細胞胚胎多為近二倍體、近單倍體,其染色體核型紊亂。 (3) ICSI-1PN胚胎多為單倍體。 (4) IVF-3PN及ICSI-1PN胚胎均不能用于移植。
[Abstract]:Aim: to analyze the chromosome composition of abnormal fertilized eggs and embryos derived from abnormal fertilized eggs during conventional in vitro fertilization and embryo transfer (in-vitro fertilization and embryo transfer, IVF-ET) by single-cell chromosome preparation technique. Methods: 75 abnormal fertilized eggs and 277 abnormal fertilized eggs were collected 24 hours after fertilization, including 71 IVF-derived tripronuclear (tripronuclear, 3PN) fertilized eggs, 38 2-cell embryos and 239 2-cell embryos. Four monopronucleus, 1PN embryos derived from intracytoplasmic sperm microinjection of (intracytoplasmic sperm injection,ICSI were obtained. Colchicine (final concentration 0.10 ug/ml) was co-cultured with colchicine for 14 hours for 20 hours. The clear zone was removed by Tyrode' 's solution. The embryos were dispersed into blastomeres. After 20 minutes of hypotonic solution of 0.9% sodium citrate for 30 minutes, gradient fixation and trypsin banding were observed. Giemsa staining and automatic karyotype analysis system (Auto-System K) were used for karyotype analysis and photography. Results: (1) chromosome preparation of IVF-3PN fertilized eggs and embryos: mitotic index and successful rate of chromosome preparation of 3PN fertilized eggs were 67.6% (48 / 71), respectively. The mitotic index and the success rate of chromosome preparation were 38.2% (29 / 76) and 38.2% (29 / 76), respectively. The mitotic index and chromosome preparation success rate of the 2-cell embryos were the lowest, 16.2% (149 / 920) and 8.2% (75 / 920), respectively. (2) chromosome composition of 3PN fertilized eggs and embryos: the triploid rate of 3PN fertilized eggs was 75.7% (28 / 37), that of 2-cell embryos was 70.8% (17 / 24), and the karyotype of 2-cell embryos was abnormal. Most of them were near diploid (56%), near haploid (32%) and triploid rate was only 9.3% (7 / 75). (3) chromosome composition of ICSI-1PN embryos: after 4 ICSI-1PN embryos were dispersed, 29 cells were obtained, of which 6 cells (from 3 embryos) could be counted, and all of them were haploid (5 cells were 23 chromosomes). 1 the cells were 25 chromosomes. Conclusion: (1) the traditional chromosome preparation method can be used to detect the whole set of chromosomes in fertilized eggs and embryos, but the technical operation is difficult and the success rate of chromosome preparation is low. This method needs to be further improved. (2) IVF-3PN fertilized eggs and-2-cell embryos were mostly triploid, 2-cell embryos were nearly diploid and nearly haploid, and their karyotypes were disordered. (3) ICSI-1PN embryos were haploid. (4) both IVF-3PN and ICSI-1PN embryos could not be used for transplantation.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2009
【分類號】：R321

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